Dmem f12

Human neuroblastoma SH-SY5Y (ATCC CRL-2266) cells were cultured using a procedure based on Wang et al. [73 (link)]. SH-SY5Y cells were cultured in Dulbecco’s modified eagle medium (DMEM)/F12 supplemented with 10% heat-inactivated fetal bovine serum (FBS) and 1% penicillin/streptomycin (PS) (growth media) from Bovogen Biologicals (Melbourne, VIC Australia). Cells were maintained at 37 °C in a humidified incubator with 5% CO₂ and 95% relative humidity and passaged every 3 days.

Immortalized mouse OECs were generated from olfactory bulb ensheathing glia of GFP-expressing mice (C57BL/6-Tg (ACTB-EGFP)1Osb/J, Jackson Laboratory, Bar Harbor, USA [7 (link), 8 (link)]. mOEC-GFP were cultured in Dulbecco’s Modified Eagle Medium/Nutrient F-12 (DMEM/F12) supplemented with 10% fetal bovine serum (FBS, Bovogen) and 10 ng/mL gentamicin (Life Technologies). Dorsal root ganglion Schwann cells and astrocytes were isolated from S100β-DsRed transgenic mice [9 (link)]. Schwann cells and astrocytes were maintained in complete medium conditions (Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% FBS, 1% GlutaMAX™ (Life Technologies) supplement and 50 ng/mL gentamicin). Pancreatic cancer cell (BxPC-3) and Neural stem cells (ReNcell VM) were maintained in DMEM supplemented with 10% FBS and 10 ng/mL gentamicin and in DMEM/F12 supplemented with 10% fetal bovine serum, 1% N-2 Supplement (Life Technologies) and 10 ng/mL gentamicin, respectively. All the cells were maintained in a humidified incubator at 37 °C and 5% CO2.

Human neuroblastoma SH-SY5Y (ATCC CRL-2266) cells were maintained at 37℃ in a humidified incubator with 5% CO₂ and 95% relative humidity. SH-SY5Y cells were cultured in Dulbecco’s modified eagle medium (DMEM)/F12 supplemented with 10% of heat-inactivated fetal bovine serum (FBS) and 1% penicillin–streptomycin from Bovogen Biologicals (Victoria, Australia). SH-SY5Y cells were differentiated, as described previously [28 (link)]. Briefly, cells were seeded in MaxGel ECM (E0282, Sigma-Aldrich, Sydney)–coated 96-well plates with media containing 10 µM retinoic acid (RA, Sigma-Aldrich, Sydney) in the dark. Cells were observed for neurite outgrowth using the IncuCyte ZOOM live-cell imaging system (Essen BioScience, UK). Neurite integrity was quantified using the automated NeuroTrack module of the IncuCyte ZOOM system for live-cell analysis. Furthermore, differentiated cells were exposed to various concentrations of CBD (0.1, 0.5, 1, 2.5, 5, 10 µM) in 0.1% DMSO and incubated for 24, 48, and 72 h.